1. Field of the Invention
The present invention relates to an inactivated vaccine against infectious disease caused by a group of Japanese encephalitis viruses of Flavivirus, and a diagnostic agent thereof. In particular, the present invention relates to an inactivated vaccine against Japanese encephalitis, a reinforced immunogen or antigen which is outstanding and useful as an active ingredient of the vaccine, and a method for producing the immunogen or antigen.
2. Description of the Related Art
Hereinafter, as a representative example of infectious disease caused by a group of Japanese encephalitis viruses, Japanese encephalitis will be illustrated, and a vaccine against it will be described. The first Japanese encephalitis vaccine was put into practical use in 1954. This vaccine contains, as its active ingredient, an antigen prepared from virus cultured in mouse brain. The purity of such a vaccine is low, and such a vaccine contains a lot of contamination, so that it might induce an allergic neurological disorder in the central nervous system. Thereafter, an improved high-purity vaccine obtained by the combination of alcohol sedimentation, treatment with protamine sulfate, ultra-centrifugation, and the like was put into practical use in 1965. Thus, the quality of the vaccine was remarkably improved. Such a vaccine and a production technique therefor have been utilized up to now (“Vaccine Handbook”, pp. 103-113, Researcher's Associates, the National Institute of Health (Japan), Maruzen (Tokyo) 1996). On the other hand, there were attempts to develop an inactivated vaccine obtained without using mouse brain. More specifically, the Committee on Japanese Encephalitis Vaccine was established in 1965, and they developed a vaccine using primary cell tissue culture. However, in terms of production cost, it was practically impossible to obtain the large amount of primary cell cultures required for large-scale production of inactivated vaccine antigens. Such a vaccine was not put into practical use, because at that time, only primary culture cells were approved for production of vaccines, and the use of a passage cell line was considered to be dangerous and was not permitted. Regarding a Japanese encephalitis live vaccine obtained using tissue culture, a live vaccine using, as its active ingredient, an attenuated virus grown in primary culture of hamster renal cells in China was put into practical use in China at around 1994. However, effectiveness and safety of the live vaccine have not been confirmed, and its use in various countries other than China, is not known. Furthermore, various Japanese encephalitis vaccines obtained by recombinant gene techniques (e.g., the second generation vaccine using an envelope (E) protein antigen, recombinant virus, or the like) have been reported since about 1986. However, all of these vaccines are in the experimental stage or a pre-clinical trial stage, and they have not been put into practical use (“Vaccine”, 2nd ed., pp. 671-713, S. A. Plotokin and E. A. Mortimer, W. B. Sauders Co. 1994; The Jordan Report, pp. 26-27, 1998).
Furthermore, regarding a technique of using a cell line for large-scale production of antigens or immunogens for an inactivated vaccine, for example, it is known to use a Vero cell for large-scale production of virus antigens used in vaccines against poliomyelitis (U.S. Pat. No. 4,525,349), rabies (U.S. Pat. No. 4,664,912), Hepatitis A (U.S. Pat. No. 4,783,407), and tick-borne encephalitis (U.S. Pat. No. 5,719,051), and the like. Among these, it is well-known that the former two have already been put into practical use. However, regarding the latter two, the safety and effectiveness as a vaccine of each antigen produced on a large scale have not been confirmed, and have not been put into practical use.
An active ingredient of a conventional Japanese encephalitis vaccine is inactivated particles of Japanese encephalitis virus grown in mouse brain. A large number of mice, measures against biohazard of infected animals, and the like are required for large-scale production of antigens for such a vaccine, which results in a high production cost. Furthermore, contamination into the product of adverse components derived from mouse brain (e.g., a basic protein which causes demyelination), and/or contamination of virus from a mouse, and the like, are always likely to be a factor. Therefore, purification steps and quality control become diverse and complicated. In addition, recently, it is difficult to obtain a large number of mice for production of vaccines, which becomes an obstacle to planned vaccine production. Furthermore, a conventional technique which sacrifices mice is becoming undesirable in view of animal protection and religion.